BMSCs-Derived Exosomes Ameliorate Pain Via Abrogation of Aberrant Nerve Invasion in Subchondral Bone in Lumbar Facet Joint Osteoarthritis.

Lumbar facet joint osteoarthritis (LFJ OA) is regarded as one of the common causes of low back pain (LBP). The pathogenesis and underlying mechanism of this disease are largely unknown, there is still no effective disease-modifying therapy. This study aims to investigate the efficacy of exosomes derived from bone marrow mesenchymal stem cells (BMSCs) on the pathogenesis and behavioral signs of LBP in the LFJ OA mouse model. The pathogenetic change in cartilage and aberrant nerve invasion in the subchondral bone of LFJ in a mouse model after treatment with BMSC-exosomes was evaluated. BMSC-exosomes could relieve pain via abrogation of aberrant CGRP-positive nerve and abnormal H-type vessel formation in the subchondral bone of LFJ. Moreover, BMSC-exosomes attenuated cartilage degeneration and inhibited tartrate-resistant acid phosphatase expression and RANKL-RANK-TRAF6 signaling activation to facilitate subchondral bone remodeling. These results indicated that BMSC-exosomes could relive behavioral signs of LBP and pathological processes in LFJ OA. BMSC-exosomes have a prominent protective effect and might be a potential therapeutic option for the treatment of LFJ OA causing LBP. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:670-679, 2020.

Acetabular labrum of hip joint in osteoarthritis: A qualitative original study and short review of the literature.

PURPOSE: Histological architecture of normal acetabular labrum regarding free nerve endings (FNEs) and mechanoreceptors (MRs) has been satisfactorily described in the literature. However, the presence of FNEs and MRs in acetabular labrum of hip joint has been analyzed only once in patients with osteoarthritis (OA). Aim of this article is to report histological distribution pattern of FNEs and MRs in acetabular labrum of patients with severe OA, at the same time conducting a comparison with normal acetabular labrum described in the literature. METHODS: Seven patients with severe hip OA were enrolled in this study. Patient selection was assisted by the utilization of specific clinical scales delineated by the American College of Rheumatology. After successful total hip arthroplasty, tissue samples of acetabular labra of seven patients were histologically processed and stained with the gold standard chloride method, which was subsequently examined under a compound microscope. RESULTS: FNEs and MRs constituted the major histological structures. Identified MRs included Pacini corpuscles, Ruffini corpuscles, and Golgi-Mazzoni corpuscles. The presence of FNEs was predominant in the middle part of the acetabular labrum, featuring a remarkable decrease in peripheral parts. In contrast, MRs were detected basically in peripheral parts and less in the middle part. CONCLUSIONS: Differentiation of the distribution pattern of MRs and FNEs in acetabular labrum of hip joint is remarkable between normal patients and patients with severe OA. The abundance of FNEs in the middle part of the pathologic labrum is mainly responsible for the observed discrimination. A "conversion" of MRs to FNEs may occur during OA progression, modulating therefore this pattern as well as the upcoming clinical manifestations.

Arthroscopic patelloplasty and circumpatellar denervation for the treatment of patellofemoral osteoarthritis.

BACKGROUND: Patellofemoral osteoarthritis commonly occurs in older people, often resulting in anterior knee pain and severely reduced quality of life. The aim was to examine the effectiveness of arthroscopic patelloplasty and circumpatellar denervation for the treatment of patellofemoral osteoarthritis (PFOA). METHODS: A total of 156 PFOA patients (62 males, 94 females; ages 45-81 years, mean 66 years) treated in our department between September 2012 and March 2013 were involved in this study. Clinical manifestations included recurrent swelling and pain in the knee joint and aggravated pain upon ascending/descending stairs, squatting down, or standing up. PFOA was treated with arthroscopic patelloplasty and circumpatellar denervation. The therapeutic effects before and after surgery were statistically evaluated using Lysholm and Kujala scores. The therapeutic effects were graded by classification of the degree of cartilage defect. RESULTS: A total of 149 cases were successfully followed up for 14.8 months, on average. The incisions healed well, and no complications occurred. After surgery, the average Lysholm score improved from 73.29 to 80.93, and the average Kujala score improved from 68.34 to 76.48. This procedure was highly effective for patients with cartilage defects I-III but not for patients with cartilage defect IV. CONCLUSIONS: For PFOA patients, this procedure is effective for significantly relieving anterior knee pain, improving knee joint function and quality of life, and deferring arthritic progression.

Characterisation and classification of the neural anatomy in the human hip joint.

Hip arthroscopy remains a useful surgical intervention for labral injuries. The literature has predominantly focused on structural and vascular considerations of the hip joint, with few studies examining the neurohistology of the surrounding periarticular tissues. We mapped and identified the periarticular neural anatomy, to identify the presence of sensory nerve fibres and mechanoreceptors within the hip joint. Eight human cadaveric hips were dissected into a total of ten specimens per hip. Histological staining was used to identify neural structures taken from the superolateral, anterior, inferior, and posterior positions of the hip joint. The frozen sections were analyzed by light microscopy to calculate relative concentrations of mean neural fibres per high power field (mnf/hpf). Neural end organs were found in the hip capsule, acetabular labrum, ligamentum teres and transverse acetabular ligament. The highest levels of mechanoreceptors were found in the superolateral aspect of the hip capsule (9.6 mnf/hpf). The labrum showed highest levels of sensory fibres (3.4 mnf/hpf) and mechanoreceptors (4.3mnf/hpf) within the anterior zone. Sensory fibres and mechanoreceptors densely populate the acetabular labrum, capsule and transverse acetabular ligament. The anterior zone of the labrum contained the highest relative concentration of sensory fibres, specifically Ruffini corpuscles.

Neurovascular invasion at the osteochondral junction and in osteophytes in osteoarthritis.

BACKGROUND: Normal adult articular cartilage is thought to be avascular and aneural. OBJECTIVE: To describe neurovascular structures at the osteochondral junction and in osteophytes in tibiofemoral osteoarthritis (OA) displaying a range of severity of cartilage changes. METHODS: Articular surfaces were obtained from 40 patients at total knee joint replacement surgery for tibiofemoral OA (TKR) and seven patients post mortem (PM). Antibodies directed against CD34 (vascular endothelium), protein gene product 9.5 (pan-neuronal marker), substance P and calcitonin gene-related peptide (sensory nerves) and C-flanking peptide of neuropeptide Y (sympathetic nerves) were used to localise blood vessels and nerves by immunohistochemistry. Severity of OA cartilage changes was graded histologically. RESULTS: TKR and PM samples displayed a range of OA cartilage changes including tidemark breaching by vascular channels. Sympathetic and sensory nerves were both present within vascular channels in the articular cartilage, in both mild and severe OA. Perivascular and free nerve fibres, and nerve trunks were observed within the subchondral bone marrow and within the marrow cavities of osteophytes. Sensory and sympathetic nerves displayed similar distributions in each region studied. CONCLUSION: Vascularisation and the associated innervation of articular cartilage may contribute to tibiofemoral pain in OA across a wide range of structural disease severity.

Decomposition of a parallel cascade model of ankle stiffness using subspace methods.

Joint stiffness, defined as the relation between the angular position of a joint and the torque acting about it, can be used to describe the dynamical behavior of the human ankle during posture and movement. Joint stiffness can be separated into intrinsic stiffness and reflex stiffness, which are modeled as a linear system and a Hammerstein system, respectively. A two-pathway parallel cascade model, with the intrinsic stiffness on one pathway and the reflex stiffness on the other, can be used to describe the joint stiffness. In this paper, we present a new method to separate the torque from each pathway from the total torque measurement. A subspace based system identification method is used to estimate the dynamics of each pathway directly from measured data without iteration. Simulation studies demonstrate that the method produces accurate results without the need of iteration.

Identification of time-varying intrinsic and reflex joint stiffness.

We have developed a time-varying, parallel- cascade system identification algorithm to separate joint stiffness into intrinsic and reflex components at each point in time throughout rapid movements. The components are identified using an iterative algorithm in which intrinsic and reflex dynamics are identified using separate time-varying (TV) techniques based on ensemble methods. An ensemble of input-output records having the same TV behavior is acquired and used to identify the system dynamics as impulse response functions at time increments corresponding to the sampling interval. Simulation studies showed that the time-varying, parallel-cascade algorithm performed well under realistic conditions with 99.9% VAF between simulated and predicted torque. To evaluate the performance of the algorithm under realistic conditions we applied it to an ensemble of experimental data acquired under stationary conditions. Results demonstrated that the TV estimates converged to those of the established time-invariant algorithm and allowed us to determine how variance of the TV estimates varied with the number of realizations in the ensemble.

Real-time estimation of intrinsic and reflex stiffness.

Joint stiffness is defined as the dynamic relationship between the position of the joint and torque acting about it. Joint stiffness is composed of two components: intrinsic and reflex stiffness. Measuring the two stiffness components cannot be done simply because the two components appear and change together. A number of approaches have been used to estimate the components, but all those approaches are inherently off-line. We have developed novel algorithm that separates and estimates the two components in real-time. Intrinsic stiffness was estimated by finding the cross-correlations between the position, its derivatives and the torque. Reflex stiffness was estimated by finding the IRF between the half-wave rectified velocity and the estimated reflex torque. A novel position perturbation, consisting of pseudo random series of pulses of different lengths, was used to eliminate covariance of intrinsic and reflex stiffness estimates. Using simulated data, the real-time estimates were shown to be estimated accurately. The real-time estimation algorithm was validated by comparing the real-time estimates with estimates generated by the parallel-cascade identification, an established off-line intrinsic and reflex stiffness identification algorithm, using simulated and experimental data. The estimates produced by the two algorithms were in agreement for both simulated and experimental data.

[Localization of the neurokinin 1 receptor in hip joints of patients with painful osteoarthritis]. / Die Lokalisation des Neurokinin 1-Rezeptors im Hüftgelenk von Patienten mit schmerzhafter Osteoarthrose.

INTRODUCTION: Recent studies on osteoarthritis have focused on nociceptive substance P (SP) containing afferent nerve fibres. The effects of SP are known to be mainly mediated by the tachykinin receptor neurokinin 1 (NK1-R). AIM: The aim of the present study was to demonstrate the NK1-R in human joint tissues. METHODS: The hip joint capsule of three patients with painful hip osteoarthritis (Group 1), three patients with femoral neck fracture showing no cartilage destruction (Group 2, controls) and the soft tissue of the fossa acetabuli of Group 1 were resected during hip arthroplasty implantation. The tissue samples were cut into small blocks and immersion-fixed in Zamboni's fixative. The specimens were frozen, cut into 50 microm sections and immunostained using a standard immunohistochemical staining protocol. RESULTS: In Groups 1 and 2 the NK1-receptor was localised in the wall of venous vessels, on Schwann cells of nerve bundles and on nerve fibres. In the osteoarthritis group the staining pattern was similar but the number of NK1-bearing cell structures seemed to be enhanced. CONCLUSIONS: The present study provides the first evidence of NK1-R in the human hip joint. In patients with painful osteoarthritis the density of NK1-R-positive cell structures seemed to be increased. The localisation of the NK1 receptor on different cell types suggests multiple effects of SP in normal and osteoarthritic joints.

Mechanoreceptors and nerve endings of the triangular fibrocartilage in the human wrist.

PURPOSE: To increase our understanding of the mechanism of pain and the sensation of wrist instability by studying the distribution of the mechanoreceptors in the triangular fibrocartilage (TFC). METHODS: The distribution and density of the nerve endings were investigated in 34 TFC specimens obtained from human cadavers. We studied the dorsal, palmar, ulnar, radial, and central areas after staining by a modified gold chloride technique. RESULTS: The free nerve endings, responsible for sensing pain, predominate in the ulnar and dorsal areas. The Vater-Pacini corpuscles predominate in the radial and dorsal area, promoting perception of the onset or cessation of movement and mechanical stress change. The Golgi-Mazzoni corpuscles were more frequent in the ulnar and ventral areas, linking these areas to function of slow adaptation and sensation of extreme movements. The proprioceptive function receptors were found in all areas of TFC because Ruffini corpuscles have homogeneous distribution in this fibrocartilaginous tissue. CONCLUSIONS: Our results showed that the nerve endings were distributed at the periphery of TFC and showed different concentrations of each type of mechanoreceptors per topographic area, suggesting that they play specific roles in the proprioceptive and nociceptive reflexes of the wrist.

Sensitisation of group III articular afferents to mechanical stimuli by substance P.

OBJECTIVE: To examine the excitatory and sensitising effects of substance P (SP) on articular afferents in normal and acutely inflamed cat knee joints. MATERIALS AND METHODS: In anesthetised cats recordings were made from 15 group III (conduction velocity 2.5-20 m/s) and 25 group IV afferent units (conduction velocity < 2.5 m/s) of the medial articular nerve of normal and acutely inflamed knee joints. SP (10 and 100 microg) was administered close-arterially. RESULTS: SP at doses of 10 microg and 100 microg activated less than 50% of both group III and group IV units. The proportion of SP-positive units was significantly higher in inflamed (10 of 21) than in normal joints (2 of 18). SP induced activity in initially silent units or increased ongoing activity after latencies varying from 2 s to 5 min. The SP-evoked activity had an irregular pattern, a variable duration, and was not related to the dose injected. Bolus injections of SP (100 microg) sensitised group III articular afferents but not group IV units to noxious movements of the joint, regardless whether the units were from normal or acutely inflamed joints. The responses to innocuous movements were not influenced by SP. Group III units, initially not activated by any movement, displayed vigorous discharges to noxious movements after close-arterial SP. In 3 group III units tested, the SP-induced augmentation of responses to noxious movements was not mimicked by close-arterial injection of histamine (3.3 microg). CONCLUSIONS: It is concluded that SP contributes to the sensitisation of a subpopulation of high-threshold articular afferents. Thus this neuronal mediator released peripherally in response to an injury or acute inflammation causes considerable changes in the mechanosensitivity of this subpopulation of nociceptive joint afferents.

The innervation of the triangular fibrocartilage complex: nitric acid maceration rediscovered.

Injury to the triangular fibrocartilage complex (TFCC) is frequently implicated in the etiology of ulnar-sided wrist pain. This study examines the nervous anatomy of the TFCC using a nitric acid maceration technique and attempts to correlate this information with known tear patterns. Ten fresh frozen cadaveric specimens were studied in detail. Gross dissection of each upper-extremity specimen included removal of all flexor and extensor tendons. After identification and labeling with permanent color of the ulnar nerve, dorsal sensory branch of the ulnar nerve, posterior interosseous nerve, anterior interosseous nerve, and median nerve, an en bloc excision of the distal radioulnar region was performed. Digestion of the soft tissue was performed with nitric acid at sequential concentrations of 50% and 33% for 9 of 10 specimens. The digestion was halted by immersing the specimen in a mixture of 10% formaldehyde and 1% glycerine. After removal of bone, the specimens were fixed in paraffin, sectioned, and stained with hematoxylin and eosin. Nine of the 10 specimens were studied microscopically to determine the contribution of the grossly identified nerves to each zone of the triangular fibrocartilage complex as defined by Palmer's classification of acute TFCC tears. The anterior interosseous, median, and superficial radial nerves did not contribute to the innervation of the TFCC. The intraarticular course of the peripheral nerves could not be defined in the one specimen that was not digested with nitric acid. Nitric acid maceration is a rediscovered technique for identifying the nervous anatomy of soft tissues. The study showed that the triangular fibrocartilage complex is innervated by branches of the posterior interosseous, ulnar, and dorsal sensory ulnar nerves in a fairly consistent manner. Improved treatment of TFCC tears may result from an enhanced understanding of the supporting structures' innervation and mechanical function.

Neurophysiological basis for neurogenic-mediated articular cartilage anabolism alteration.

This study was designed to investigate the pathways involved in neurogenic-mediated articular cartilage damage triggered by a nonsystemic distant subcutaneous or intra-articular inflammation. The cartilage damage was assessed 24 h after subcutaneous or intra-articular complete Freund's adjuvant (CFA) injection measuring patellar proteoglycan (PG) synthesis (ex vivo [Na(2)(35)SO(4)] incorporation) in 96 Wistar rats. Unilateral subcutaneous or intra-articular injection of CFA induced significant decrease (25-29%) in PG synthesis in both patellae. Chronic administration of capsaicin (50 mg. kg(-1). day(-1) during 4 days), which blunted the normal response of C fiber stimulation, prevented the bilateral significant decrease in cartilage synthesis. Similarly, intrathecal injection of MK-801 (10 nmol/day during 5 days), which blocked the glutamatergic synaptic transmission at the dorsal horn of signal originating in primary afferent C fibers, eliminated the CFA-induced PG synthesis decrease in both patellae. Chemical sympathectomy, induced by guanethidine (12.5 mg. kg(-1). day(-1) during 6 wk), also prevented PG synthesis alteration. Finally, compression of the spinal cord at the T3-T5 level had a similar protective effect on the reduction of [Na(2)(35)SO(4)] incorporation. It is concluded that the signal that triggers articular cartilage synthesis damage induced by a distant local inflammation 1) is transmitted through the afferent C fibers, 2) makes glutamatergic synaptic connections with the preganglionic neurons of the sympathetic system, and 3) involves spinal and supraspinal pathways.

Galanin influences the mechanosensitivity of sensory endings in the rat knee joint.

The aim of the present study was to examine the effect of galanin on group III and IV afferent nerve fibres (n = 53) innervating normal and acutely inflamed knee joints in rats. They responded to local mechanical stimulation, movements of the joint and i.a. injections of KCl close to the joint. Single i.a. bolus injections of galanin (0.1 mM, 0.2 mL) caused no direct responses of the units. In normal and acutely inflamed joints, about half of the units did not change the responses to knee joint rotation. A significant reduction of the responses to noxious movements was found in approximately 40% of the units reaching a mean value of 57% in normal joints and 70% in inflamed joints compared with control movements. In approximately 10% the responses increased to 143% in normal joints and 120% in inflamed joints. Injection of a galanin receptor antagonist (M35) doubled the responses to noxious movements in 36% of the units in normal joints and reduced it in 18% to 86% of the control movements, indicating a tonic release and influence on the mechanosensitivity of a proportion of primary afferents by galanin. In conclusion, these data further support the hypothesis that the mechanosensitivity of fine afferent nerve fibres is regulated by a mixture of different substances being released into the innervated tissue. Besides the action of several pro-inflammatory peptides there seems to exist a tonic inhibitory system.

Retrograde tracing and neuropeptide immunohistochemistry of sensory neurones projecting to the cartilaginous distal femoral epiphysis of young rats.

Although cartilage is considered to be devoid of innervation, axons occur in the perichondrium and during development in cartilage canals, thereby having a relatively close spatial relationship to chondroblasts and chondrocytes. The present study locates the source of the sensory innervation of the femoral cartilaginous epiphyses of young rats and investigates whether the neuropeptide calcitonin gene-related peptide (CGRP) can influence chondrocytes. Retrograde tracing from the distal femoral epiphysis of young rats with Fast Blue (FB) showed labelled neuronal profiles in the L2-L5 dorsal root ganglia. Sample countings indicated that 50% of the FB-labelled neuronal profiles were located at the L3 level and 25% at the L4 level. The labelled neurones had diameters of 15-40 microm, with a peak at 25-30 microm. Immunohistochemistry showed that about 50% of the FB-labelled profiles contained CGRP. Together with the finding that CGRP influences bone cells to generate the second messenger cAMP, this result suggested the hypothesis that chondrocytes might be similarly influenced by CGRP. However, stimulation of cartilage slices with CGRP in vitro followed by an assay of the cAMP content did not provide support for this hypothesis. We conclude that primary sensory neurones containing CGRP project to the perichondrium and to cartilage canals of growing cartilage, and that exogenous CGRP does not elevate the cAMP content of cartilage slices in vitro.

[Deterioration of sensory joint innervation as an enabling factor for development of arthrosis. An animal experiment study of the rat model]. / Störung sensibler Gelenkinnervation als begünstigender Faktor für die Arthroseentstehung. Eine tierexperimentelle Untersuchung am Rattenmodell.

In the present study we investigated the influence of an altered sensible joint innervation on the development of knee osteoarthritis in a wistar rat model of osteoarthritis. Capsaicin (8-methyl-N-vanillyl-6-noneamide) mediated partial sensible knee joint denervation was performed in a group of 16 male wistar rats. Twelve rats without alterations of the sensible knee joint innervation served as controls. In both groups, half of the rats underwent strenuous running exercises (total running load of 20 km) in a running wheel by intracranial self-stimulation, while the other half did not have any running load. In rats without running, there were no histological sings of knee osteoarthritis according to the Mankin score. In contrast, in rats running a total of 20 km significant osteoarthritis changes were observed. Hereby, in rats without altered sensible knee joint innervation, osteoarthritis was mostly classified as mild or moderate, while severe osteoarthritis was the predominant finding in the knee joints of the rats with partial sensible knee joint denervation. In conclusion, our study gives strong evidence for the hypothesis that an altered sensible joint innervation works as a contributing factor in the development of osteoarthritis.

Gold chloride technique to study articular innervation. A protocol validated through computer-assisted colorimetry.

We studied variations in gold chloride techniques to elicit neural elements within articular samples, after "in toto" staining. These techniques attempt the differentiation of neural and vascular structures. Major changes in differential staining were observed when the gold chloride concentration was empirically modified. After the rest of the technique was standardized, we selected three gold chloride solutions to perform quantitative color experiments: 1%, 0.75%, and 0.5%. Significant sections of the same thickness were acquired with a digital camera to perform computer-assisted colorimetry. Color was measured through RGB (red-green-blue) channels in vessels, nerves, and background connective tissue as an internal control. By means of multivariate regression analysis, we compared differences in color measurements after 1%, 0.75% and 0.5% gold chloride preparation. Statistically significant coefficients confirmed that red color signals in vessels after the 0.75% and the 0.5% solution were both less intense than after the 1% preparation. Green and blue signals in vessels were also significantly less intense after the 0.5% protocol than after using the 1% solution. Red color signals in nerves between the 1% and the 0.75% preparation protocols were more intense and not significantly different, while the 0.5% preparation produced significantly less intense red signals in nerves. Non-significant differences were observed in green or blue signals from nerves after any protocol. We concluded that the 0.75% gold chloride solution protocol produced more intense red signals in nerves and less intense red signals in vessels. This was the most discriminant protocol in our series, based on color signals.

A study of mechanoreceptors in fibrocartilage masses in the defect of pars interarticularis.

We investigated the origin of low back pain associated with lumbar spondylolysis and spondylolytic spondylolisthesis by removing fibrocartilage masses from the lytic sites in symptomatic patients and staining the masses by the Gairns gold chloride method to examine mechanoreceptors. The fibrocartilage masses were found to contain four types of mechanoreceptors: Pacinian corpuscles, Ruffini receptors, Golgi tendon organ-like receptors, and free nerve endings. All of these mechanoreceptors were present at the periphery of the specimens, and Ruffini receptors and free nerve endings were abundant. Some mechanoreceptors had a slightly atypical structure, in addition to those with typical morphology. Comparison with mechanoreceptors in normal lumbar facet joint capsules showed that there were more mechanoreceptors in the fibrocartilage masses and a greater proportion of atypical structures at lytic sites. The presence of mechanoreceptors at lytic sites suggests that the fibrocartilage masses are not simply scar tissue filling the defect. Rather, these masses also appear to play a protective role by sensing instability via mechanoreceptors and transmitting this information as pain, while at the same time acting as ligament-like tissue that connects and stabilizes the separated vertebral arches.

Morphology and distribution of nerve endings in the human triangular fibrocartilage complex.

We studied the morphology and distribution of nerve endings in the human triangular fibrocartilage complex using both silver staining and immunohistochemical staining using a protein specific to nerve fibres. Free nerve endings were found in the ulnar side of the triangular fibrocartilage complex, especially in the ulnar collateral ligament, meniscus homologue and the adjacent collagen fibre area of the peripheral part of the ulnar side of the articular disc. Meissner's and Krause's corpuscles were observed in the ulnar collateral ligament and meniscus homologue. The fact that free nerve endings were observed in the meniscus homologue and adjacent collagen fibre area of the peripheral part of the ulnar side of the articular disc suggests that this disc may be a source of wrist pain. The presence of nerve end bulbs in the triangular fibrocartilage complex also suggests a possible role for corpuscles as mechanoreceptors.

Functional anatomy of the cartilage of the distal phalanx and digital cushion in the equine foot and a hemodynamic flow hypothesis of energy dissipation.

OBJECTIVES: To examine macro- and microscopic characteristics of cartilage of the distal phalanx (ungual cartilage [UC]) and digital cushion in the equine foot and to relate them to the foot's function of energy dissipation. ANIMALS: 85 horses and 5 foals of various breeds and ages. PROCEDURE: Feet, obtained at necropsy, were perfused with India ink (n = 30), latex (5), or polymer plastic (10). Select feet were examined histologically for tissue architecture and to identify elastic fibers. Immunochemistry to identify substance P peptides in nerves (feet from foals) and gold chloride impregnation of axons (n = 10) were performed. Feet were sectioned transversely (n = 27) or coronally (62 feet in a matched-paired study). Ungual cartilage was measured at the navicular bone. Digital cushions were examined for relative tissue composition between forefeet and hind feet. RESULTS: Ungual cartilage formed an axial projection that extended towards the midline to overlie the bars, and dorsally along the semilunar line of the distal phalanx. Ungual cartilage of forefeet was significantly larger than that of hind feet. The digital cushion was composed of fat and elastic tissues in feet with thin UC, or fibrous and fibrocartilaginous tissue and elastic tissue in feet with thicker UC. Sensory nerves and an extensive network of venovenous anastomoses were found in the UC. CONCLUSIONS AND CLINICAL RELEVANCE: Ungual cartilage and the digital cushion provide the basis for a hemodynamic flow hypothesis of energy dissipation. Maximum energy dissipation depends on proper hoof preparation and shoeing.